Transient cerebral ischemia/stroke impairs endoplasmic reticulum (ER) function, which in turn triggers translation arrest that is irreversible in vulnerable neurons. Cells cannot survive when ER function is irreversibly damaged by a severe form of stress. For functional recovery of neurons following a transient interruption of blood supply, restoration of ER function is therefore as crucial as reperfusion of the ischemic tissue. However, the precise role of ER dysfunction in the pathological process that culminates in ischemic cell death of neurons has never been established. It is of key importance to fill this gap in our knowledge because until we do, the possibility of making neurons more resistant to transient ischemia by restoring ER function will remain unexplored. The long-term goal is to understand how ER functioning can be manipulated for preventive and therapeutic purposes. The objective here is to create transgenic animals expressing the short form of ATF6, a transcription factor required to restore impaired ER function. Once created, the transgenic animals will be a pivotal platform to elucidate how restoration of ER function modulates recovery of neurons from the severe metabolic stress induced by ischemia. The central hypothesis is that restoration of ER function is essential for recovery of neurons from transient ischemia. This hypothesis has been formulated on the basis of data generated in our laboratories. The rationale for the proposed research is that once we have established the role of impaired ER function in ischemia-induced cell death, new and innovative approaches can most probably be developed to prevent or treat a variety of diseases associated with impaired ER function. Guided by strong preliminary data, this hypothesis will be tested by pursuing the following specific aim: Create transgenic mice expressing ATF6 after crossbreeding with Cre transgenic animals. The ATF6xCre mice will express ATF6 that is retained in the cytoplasm and requires induction by tamoxifen for nuclear translocation and thus activation of the expression of ATF6-dependent genes. After tamoxifen activation, animals will be subjected to transient focal cerebral ischemia to investigate the effects of induced expression of ATF6 and ATF6-dependent genes on size of infarcts. The approach is innovative, because it focuses for the first time on manipulating ER function of neurons for therapeutic purposes in stroke. The proposed research is significant, because it is expected to lead to the development of new strategies for preventive and therapeutic interventions to render cells more resistant to a transient episode of cerebral ischemia/stroke. Furthermore, the ATF6 transgenic animals will provide a promising tool for future studies to investigate the role of ER dysfunction in a large variety of disorders of high clinical relevance, after cross- breeding with Cre transgenic animals to specifically express ATF6 in the organ/cell type under investigation. PUBLIC HEALTH RELEVANCE: The proposed research is relevant to public health because elucidating the role of endoplasmic reticulum dysfunction in ischemia/stroke-induced neuronal cell death is ultimately expected to help identifying new targets for therapeutic intervention. Thus, the proposed research is relevant to the part of NIH's mission to foster fundamental creative discoveries, innovative research strategies, and their applications as a basis to advance significantly the Nation's capacity to protect and improve health.